SEPON, a Selection and Evaluation Pipeline for OligoNucleotides based on ESTs with a non-target Tm algorithm for...

Area of Science:

• Genomics
• Bioinformatics
• Molecular Biology

Background:

• Microarray gene-expression profiling requires high-quality oligonucleotide sequences.
• Non-annotated genomes present challenges in selecting specific oligonucleotide probes.
• Cross-hybridization can lead to inaccurate gene expression data.

Purpose of the Study:

• To develop a computational pipeline for selecting optimal oligonucleotide sequences from expressed sequence tags of non-annotated genomes.
• To implement a non-target melting temperature (T(m)) algorithm to reduce cross-hybridization.
• To provide a user-defined filtering and ranking system for oligonucleotide selection.

Main Methods:

• The Selection and Evaluation Pipeline for OligoNucleotide (SEPON) was developed.
• An algorithm was created to estimate the T(m) of oligonucleotide hybridization to non-specific targets (non-target T(m)).
• Oligonucleotides with non-target T(m) above a user-defined threshold were discarded.
• Filtering criteria included predicted exon location, 3' distance, GC content, secondary structure T(m), and non-target T(m).

Main Results:

• SEPON generates n-mer oligonucleotide sequences suitable for microarray gene-expression profiling.
• The non-target T(m) algorithm effectively reduces potential cross-hybridization.
• The pipeline ranks oligonucleotides based on multiple user-defined parameters for optimal selection.

Conclusions:

• SEPON provides an effective method for selecting specific oligonucleotide sequences from non-annotated genomes.
• The tool enhances the accuracy of microarray gene-expression profiling by minimizing cross-hybridization.
• SEPON offers a flexible and customizable approach to oligonucleotide design.